Rapid GAL Gene Switch of Saccharomyces cerevisiae Depends on Nuclear Gal3, Not Nucleocytoplasmic Trafficking of Gal3 and Gal80
Rapid GAL Gene Switch of Saccharomyces cerevisiae Depends on Nuclear Gal3, Not Nucleocytoplasmic Trafficking of Gal3 and Gal80
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DOI：
10.1534/genetics.111.131839
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发表时间：
2011-11-01
期刊：
影响因子：
3.3
通讯作者：
Hopper, James E.
中科院分区：
文献类型：
--
作者：
Egriboz, Onur;Jiang, Fenglei;Hopper, James E.
酵母转录激活因子Gal 4即使在半乳糖不存在的情况下也定位于UASGAL位点，但由于与Gal 80蛋白的结合而不能激活转录。在半乳糖添加后4 min，Gal 4激活的基因转录增强。已经确定，这种快速诱导是通过半乳糖触发的Gal 80和Gal 3蛋白之间的结合而产生的，该结合降低了Gal 80和Gal 4的结合。这是如何发生的机制尚不清楚。关于Gal 3和Gal 80的核质分布和运输的可能作用以及在细胞中最初的Gal 3-Gal 80缔合发生的地方，存在着显著不同的假说。在这里，我们测试了两个相互矛盾的假设，通过评估的亚细胞分布和动力学的Gal 3和Gal 80的诱导动力学。我们确定Gal 80和Gal 3的核质运输速率相对于诱导速率是缓慢的。我们发现，耗尽核池的Gal 3减慢的诱导动力学。因此，核Gal 3对于快速诱导至关重要。光漂白后的光漂白恢复实验提供的数据表明Gal 80-Gal 4复合物在不存在半乳糖的情况下表现出动力学稳定性。最后，我们检测Gal 3在UASGAL只有当Gal 80共价连接到DNA结合结构域。总之，这些新的发现使我们提出，一个短暂的相互作用的Gal 3与Gal 4相关的Gal 80可以解释这个系统的快速反应。这个概念也可以解释先前的观察。
The yeast transcriptional activator Gal4 localizes to UASGAL sites even in the absence of galactose but cannot activate transcription due to an association with the Gal80 protein. By 4 min after galactose addition, Gal4-activated gene transcription ensues. It is well established that this rapid induction arises through a galactose-triggered association between the Gal80 and Gal3 proteins that decreases the association of Gal80 and Gal4. How this happens mechanistically remains unclear. Strikingly different hypotheses prevail concerning the possible roles of nucleocytoplasmic distribution and trafficking of Gal3 and Gal80 and where in the cell the initial Gal3-Gal80 association occurs. Here we tested two conflicting hypotheses by evaluating the subcellular distribution and dynamics of Gal3 and Gal80 with reference to induction kinetics. We determined that the rates of nucleocytoplasmic trafficking for both Gal80 and Gal3 are slow relative to the rate of induction. We find that depletion of the nuclear pool of Gal3 slows the induction kinetics. Thus, nuclear Gal3 is critical for rapid induction. Fluorescence-recovery-after-photobleaching experiments provided data suggesting that the Gal80-Gal4 complex exhibits kinetic stability in the absence of galactose. Finally, we detect Gal3 at the UASGAL only if Gal80 is covalently linked to the DNA-binding domain. Taken altogether, these new findings lead us to propose that a transient interaction of Gal3 with Gal4-associated Gal80 could explain the rapid response of this system. This notion could also explain earlier observations.